Cancer is a leading cause of death in the world. Despite the development of newer chemotherapeutic agents and combination chemotherapy regimens, metastatic neoplastic diseases are often resistant to therapy. The reasons for this drug-resistance are two-fold. First, specific genes may be expressed that impart drug-resistant characteristics to the neoplastic tissue. Second, tumors are often heterogeneous tissues in their sensitivity to specific chemotherapeutic agents. Treatment over time thus selects out the resistant tissue. An understanding of drug-resistance gene expression within a tumor over time thus is of importance in developing appropriate treatment regimens for the patient. Current methods for evaluating the drug-resistance phenotype of a patient's tumor require the analysis of a tissue specimen obtained by an invasive biopsy of the tumor. The invasive nature of these biopsies often precludes the serial longitudinal monitoring of drug-resistance in a given patient, and further, is prone to sampling error.
This invention relates to methods for evaluating the expression of drug-resistance genes (drug-resistance-associated genes) in neoplastic tissue without the requirement of tissue biopsy. Specifically, the invention provides for the detection and monitoring of drug-resistance gene nucleic acid, particularly ribonucleic acid (RNA) or deoxyribonucleic acid (DNA), in a bodily fluid from an animal or human. Since bodily fluids such as blood, plasma, serum, urine, saliva, cerebrospinal fluid, and effusions are more easily and readily obtainable than most tissue specimens, the invention provides a convenient method of evaluating a tumor's drug-resistance, and thereby of selecting, monitoring, or altering drug therapies such as chemotherapy. Furthermore, the invention provides a method to evaluate drug-resistance expression of the entire tumor-burden of an animal, preferably a human, thereby reducing sampling bias induced by tumor heterogeneity, as may occur during analysis of localized tumor biopsies. The invention therefore provides methods for evaluating the presence of RNA and mutated or altered or polymorphic DNA associated with drug-resistance genes in bodily fluid, particularly blood, plasma, serum, and other bodily fluid, wherein said genes include but are not limited to the multidrug resistance 1 gene (MDR-1), said gene encoding the 170 kD transport protein P-glycoprotein (Pgp), the multidrug resistance-associated protein gene (MRP) encoding a 190 kD adenosine triphosphate binding transport protein with homology to MDR-1, and further associated genes encoding the multidrug resistance proteins MRP1, MRP2, MRP3, and MRP5, the topoisomerase I gene, the topoisomerase II alpha and beta genes, genes associated with glutathione metabolism (GSH genes) including the glutathione S-transferase genes, the thymidylate synthase gene (TS), the thymidine phosphorylase gene (TP), and the dihydropyrimidine dehydrogenase gene (DPD), said gene RNAs being characterized as tumor-associated RNA or DNA herein. Co-owned and co-pending U.S. patent application Ser. No. 09/155,152, incorporated herein by reference in its entirety, detects tumor-associated RNA in bodily fluids such as blood plasma and serum, wherein said RNA detection is used for detecting, monitoring, or evaluating cancer or premalignant conditions. In the present invention, methods for detecting extracellular nucleic acids are utilized in a novel manner to determine drug-resistance gene expression in a patient. Furthermore, a novel method is described herein that enables evaluation of drug-resistance gene expression in a tumor without the need of directly obtaining tumor tissue.
There is a newly-appreciated need in the art to identify drug-resistance propensity in an animal, most preferably a human, in a safe and convenient manner by detecting in a qualitative or quantitative fashion drug-resistance gene RNA and DNA such as MDR-1 RNA, MRP RNA, associated MRP1 RNA, MRP2 RNA, MRP3 RNA, MRP5 RNA, GSH transferase RNA, TS RNA, TP RNA, DPD RNA, mutated topoisomerase I RNA or DNA, mutated topoisomerase II alpha and beta RNA and DNA, and other mutated or altered DNA in bodily fluids such as whole blood or blood plasma or serum, including DNA polymorphisms including but not limited to MDR-1 polymorphisms, GSH-associated gene polymorphisms including GSH-S transferase polymorphisms, TS polymorphisms, and MDR-1 polymorphisms. Further, there is a need in the art to evaluate the predisposition in an animal, most preferably a human, to respond favorably or unfavorably to a particular chemotherapy regimen by detecting drug-resistance gene RNA or mutated or altered or polymorphic DNA in bodily fluids such as blood plasma or serum, thereby enabling particular treatment regimens to chosen.